Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA209358

Local Sample ID:	348
Subject ID:	SU002266
Subject Type:	Human
Subject Species:	Homo sapiens
Taxonomy ID:	9606
Gender:	Male and female

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Sample Preparation:

Sampleprep ID:	SP002272
Sampleprep Summary:	Lipids were extracted using a modified biphasic separation protocol with methyl tertiary-butyl ether (MTBE)(Matyash et al., 2008). Pipetting was performed using Rainin BioClean Ultra pipette tips. In 2 ml Protein LoBind Eppendorf tubes (cat# 022431102), 40 μl of neat plasma were mixed with 260 μl ice-cold MeOH and briefly vortexed to denature proteins. Next, 40 μL of an isotopically labeled standard lipid stock that approximates the blood plasma lipid composition was added to each sample (SCIEX, cat# 5040156, LPISTDKIT-101), which was prepared according to the instruction in Lipidomics Workflow Manager (LWM). The mix was briefly vortexed. Next, 1000 μL ice-cold MTBE was added after which samples were vortexed for 10 seconds followed by shaking at 4ºC for 30 min. Phase separation was induced by adding 250 μL of HPLC grade water. Samples were subsequently vortexed for 60 seconds. Next, samples were centrifuged at 16,000 g for 5 min at room temperature. 750 μL of the upper organic (MTBE) phase were transferred to a new 2 mL tube. This MTBE lipid fraction was dried down in a nitrogen blower for about 2 hours. For storage at -20ºC, 200 μL of MeOH was added to dried-down samples.

Sampleprep ID:

SP002272

Sampleprep Summary:

Lipids were extracted using a modified biphasic separation protocol with methyl tertiary-butyl ether (MTBE)(Matyash et al., 2008). Pipetting was performed using Rainin BioClean Ultra pipette tips. In 2 ml Protein LoBind Eppendorf tubes (cat# 022431102), 40 μl of neat plasma were mixed with 260 μl ice-cold MeOH and briefly vortexed to denature proteins. Next, 40 μL of an isotopically labeled standard lipid stock that approximates the blood plasma lipid composition was added to each sample (SCIEX, cat# 5040156, LPISTDKIT-101), which was prepared according to the instruction in Lipidomics Workflow Manager (LWM). The mix was briefly vortexed. Next, 1000 μL ice-cold MTBE was added after which samples were vortexed for 10 seconds followed by shaking at 4ºC for 30 min. Phase separation was induced by adding 250 μL of HPLC grade water. Samples were subsequently vortexed for 60 seconds. Next, samples were centrifuged at 16,000 g for 5 min at room temperature. 750 μL of the upper organic (MTBE) phase were transferred to a new 2 mL tube. This MTBE lipid fraction was dried down in a nitrogen blower for about 2 hours. For storage at -20ºC, 200 μL of MeOH was added to dried-down samples.